There is growing evidence that cell-mediated immune responses, and in particular MHC class I-restricted cytotoxic T lymphocytes (CTL), are important in controlling HIV-1 replication. Thus, one major challenge for HIV-1 vaccine development is to create novel vaccine strategies that efficiently deliver antigens into the MHC class I pathway to induce strong CTL responses against a variety of epitopes. Dendritic cells (DC) are professional antigen presenting cells (APCs) that are unique in their ability to efficently process exogenous antigens for presentation by MHC class I molecules (cross-priming, cross-presentation, and to activate naive, antigen-specific T lymphocytes. Human blood DCs, the precursors of tissue and lymph node DCs, have been categorized into two discrete subsets, myeloid DCs (mDC) and plasmacytoid DCs (pDC), based on differences in lineage, phenotype, and function. Both subsets of human DCs may play a role in mediating immune responses to vaccines. We have previously shown that whole S. cerevisiae yeast were avidly internalized by both routine and human DCs in vitro, leading to DC maturation and IL-12 production. Furthermore, whole yeast were shown to possess a class I-directed adjuvanticity, and yeast recombinant for tumor and/or HIV-1 antigens were able to induce CTL and CD8+ T cell-mediated protective tumor immunity in vaccinated animals. In this proposal, we plan to investigate the specific interactions between recombinant yeast and human blood DCs, including the mechanisms of DC activation, the pathways involved in processing yeast-derived antigens, and the breadth of CTL epitopes presented. The proposed studies will not only facilitate the development of recombinant yeast as an HIV-1 vaccine, but should also promote development of novel strategies for effectively targeting exogenous vaccine antigens into the MHC class I presentation pathway of DCs in order to promote strong and broadly-directed CTL responses. [unreadable] [unreadable]